1. Field of the Invention
The present invention is directed to the amino acid sequence for cDNA encoding the tumor rejection antigen known as gp 110. More specifically, the present invention provides an amino acid sequence for murine gp110, including identification of the amino-terminal sequence as defined by automated Edman degradation sequencing. Prophylactic and therapeutic methods for using gp110 are also disclosed.
2. Background Information
T-cell mediated responses play an important role in a host""s immune response to viral infections and cancer. T-cells function by recognizing an antigen in the context of relatively short peptides. These antigenic peptides, which have been processed from exogenous and endogenous proteins, are presented on the cell""s surface in association with Class I and II MHC molecules. As a result, any cell can literally display on its cell surface an array of peptides, derived from proteins found within the cell, for continuous sampling and evaluation by various T-cell populations. For example, CD8+ cytolytic T-cells (CTL) recognize a non-covalent trimeric complex of a given MHC Class I heavy chain allele, xcex22-m, and a peptide comprised of 8 to 12 amino acid residues. Such complexes may be amino affinity purified, denatured in vitro, and the peptides isolated and sequenced. These peptides can then be used in the formation of peptide-based vaccines.
The concept of immunity against tumors is based on the fact that tumor cells carry antigens that are not detect on normal cells. Two classes of tumor antigens have been identified: tumor specific antigens (TSA) and tumor associated antigens. Tumor specific antigens are unique to tumor cells and do not occur on other cells in the body. Tumor-associated antigens are not unique to the tumor cells and instead are also expressed on normal cells under conditions that fail to induce a state of immunologic tolerance to the antigen. The expression of the antigen on the tumor may occur under conditions that enable the immune system to respond to the antigen. Tumor-associated antigens may be antigens that are expressed on normal cells during fetal development when the immune system is immature and unable to respond or they may be antigens that are normally present at extremely low levels on normal cells, but which are expressed at much higher levels on tumor cells.
In general, tumor antigens are defined by their ability to elicit a humoral and/or cell-mediated response. Although a few tumor antigens have been shown to induce the production of humoral antibodies, most induce a cell-mediated response. A class of tumor antigens has been shown to elicit cell-mediated responses in mice that result in rejection of tumors, expressing these antigens when they are transplanted into syngeneic recipients. Because of this phenomenon, these tumor antigens are referred to as tumor specific transplantation antigens (TSTA), tumor rejection antigens (TRA), or tumor-associated transplantation antigens (TATAs).
Studies of the immunogenicity of chemically induced sarcomas of inbred mice established the concept of TSA and revealed their potential role in cancer immunotherapy. Three murine TRA have been isolated from the chemically induced BALB/c Meth A sarcoma and other tumors and are believed to have restricted tumor rejection inducing activities. These three TRA are known as Mr 82,000 (p82), Mr 84,000/86,000 (HSP84/86) and Mr 96,000 (gp96). The HSP84/86 and gp96 TRA are stress-induced proteins; the HSP84/86 antigens are the murine equivalents of the Mr 90,000 heat-shock proteins, while gp96 is identical to the glucose-regulated protein GRp94.
A Meth A TRA identified as Mr 110,000 (gp110) appears to be antigenically distinct from previously identified TRA isolated from Meth A, and responsible for the highly restricted tumor rejection-inducing activity of this tumor. The gp110 family of tumor antigens is a novel set of tumor rejection antigens that appear to be responsible for the antigenic diversity of chemically-induced sarcomas and other tumor types known to express antigenically diverse TRAs as well. Based on their well-characterized functional activity as tumor rejection antigens, the translational potential of gp110 in the development of clinical peptide-based vaccines is significant.
DeLeo, et al. xe2x80x9cProperties of a Mr 110,000 Tumor Rejection Antigen of the Chemically Induced BALB/c Meth A Sarcomaxe2x80x9d, Cancer Res. 53:1602-1607 (1993) reported that it appears to be gp110, and not gp96, that has tumor rejection inducing activity.
Fassanito, et al. xe2x80x9cCharacterization of Cloned Class I MHC-restricted, CD8+ Anti-Meth A Cytotoxic T-Lymphocytes: Recognition of an Epitope Derived from the Meth A gp110 Tumor Rejection Antigenxe2x80x9d, Cancer Res. 55:124-128 (1995) reported that the determinant recognized by the cloned anti-Meth A CTL line (CTLMA-ac) is derived from Meth A gp110.
Mayordomo, et al. xe2x80x9cBone Marrow-derived Dendritic Cells Pulsed with Synthetic Tumor Peptides Elicit Protective and Therapeutic Antitumor Immunityxe2x80x9d, Nature Medicine 1:1297-1302 (1995) discuss the use of tumor peptide-pulsed dendritic cells.
Mayordomo, et al. xe2x80x9cTherapy of Murine Tumors with p53 Wild-type and Mutant Sequence Peptide-based Vaccinesxe2x80x9d, J. Exp. Med., 183:1357-1365 (1996) discuss the immunization of mice with bone-marrow derived dendritic cells prepulsed with Meth A p53 mutant peptide.
None of these articles provided the sequence of the gp110 molecule, disclosed the use of gp110 in CTL-defined peptide vaccines for the prophylactic treatment of a patient, or disclosed the use of a protein fragment, or cDNA encoding a protein fragment, comprising an amino terminal portion of gp110 for the therapeutic treatment of a patient.
The present invention is directed to the gp110 cDNA, defined as a cDNA encoding an amino acid sequence for murine gp110 molecule. This cDNA encodes a polypeptide, isolated by chromatography methods from Meth A sarcoma, comprising the N-terminal peptide of gp110 as determined by Edman sequencing, but does not include the initiator start codon for methionine. This cDNA clone also comprises the 3xe2x80x2 untranslated region of gp110. More specifically, the present invention provides the amino acid sequence of this gp110 protein fragment. Methods for using gp110 based immunotherapy in both prophylactic and therapeutic settings are also disclosed.
It is therefore an object of the present invention to provide the amino acid sequence for the gp110 cDNA encoding the murine gp110 molecule.
A further object of the present invention is to provide prophylactic resistance to tumors in a patient.
Another object of this invention is to provide peptide-based and DNA based vaccines that induce anti-peptide CTL.
A further object of the invention is to provide therapeutic relief to patients with cancers expressing gp110 and variants of it.
These and other objects of the invention will be apparent from the following disclosure and claims.